Chapter 13
Transcriptome Analysis of Regulatory Networks

13.1 Introduction 13.2 Materials 13.3.1.1 Cell Harvesting 13.3.1.2 For E.coli 13.3.2 RNA purification 13.3.2.1 Yeast 13.3.2.2 E. coli 13.3.2.3 DNaseI digestion to remove residual DNA 13.3.2.4 Quantify and check RNA quality 13.3.3 Transcriptional profiling using DNA microarrays 13.3.3.1.1 Labeling 13.3.3.1.2 Hybridization 13.3.3.1.3 Washing and scanning 13.3.3.2.1 For mRNA 13.3.3.2.2 For total RNA 13.3.3.2.3 Hybridize to GeneChip arrays, wash, and scan following manufacturer’s instructions 13.4 Data Acquisition, Anticipated Results and Interpretation 13.4.1 Acquisition of DNA microarray data 13.4.2 Normalization 13.4.3 Network Component Analysis (NCA) 13.5 Discussion and Commentary 13.6 Troubleshooting Table 13.7 Application Notes 13.8 Summary Points   References

Chapter 14
A Workflow From Time Series Gene Expression Profiling to Transcriptional Regulatory Networks

14.1		Introduction
14.2		Materials
14.3		Methods
14.3.1		Identification Of Differentially Expressed Genes
14.3.1.1		Mixed-Effects Anova Of The Normalized Time Series Data
14.3.1.2		Local False Discovery Rate Analysis
14.3.2		Robust Clustering Of Differential Gene Expression Time Series Data Using Computational Negative Control Approach
14.3.3		Transcriptional Regulatory Network Analysis Using Paint
14.3.3.1		Generation Of Paint-Compatible Input Files
14.3.3.2		Identification Of Over-Represented Transcription Factor Binding Sites
14.4		Data Acquisition, Anticipated Results, And Interpretation
14.4.1		Selection Of Number Of Clusters:
14.4.2		Paint Result Interpretation For Gene Co-Expression Clusters:
14.5		Discussion And Commentary
14.5.1		Estimation Of Non-Differentially Expressed Genes (Pi.Not Value):
14.5.2		Threshold For Local False Discovery Rate Analysis:
14.5.3		Format Of Gene Identifiers:
14.5.4		Cluster Size Issues:
14.5.5		Transfac Version Issues:
14.5.6		Annotation Redundancy In The Gene List And Multiple Promoters:
14.5.7		Reference Feasnet Selection/Generation:
14.5.8		Multiple Testing Correction In Paint:
14.6		Troubleshooting Table
14.7		Application Notes
14.8		Summary Points
		Acknowledgements
		References

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